供体肾周ad-SVF对心脏死亡来源移植肾缺血再灌注损伤的修复作用及机制

Donor after cardiac death (DCD) has contributed to the major organ source for renal transplant in China. However, during the wait for donation, recovery, storage and transplantation procedures, renal allograft suffers severe ischemia reperfusion injury (IRI), which could influence the graft function greatly. Our previous studies have confirmed that adipose stromal vascular fraction (ad-SVF) is a group of heterogeneous cells including mesenchymal stem cell (MSC) and endothelial progenitor cell (EPC), which is characterized with potent renoprotection, easy approach and no need of in vitro culture. Furthermore, the SVF derived from the perinephric adipose could express erythropoietin (EPO) specificly. However, the effects of donor perinephric adipose derived SVF on the repair of injured DCD renal allograft remain elusive. Here comes our speculation, donor perinephric adipose derived SVF might exert renoprotective functions against IRI of DCD reanl allografts. In the current project, after the establishment of rat DCD transplant model, the in vitro and in vivo experiments will be conducted with the methods such as flow cytometry for cell isolation, microPET-CT, and gene transfection, with the aim to observe the therapeutic effects of donor perinephric adipose derived SVF on the peritubular capillaries and renal tubular epithelia, and explore the potential underlying mechanisms of MSC, EPC, and EPO, which will provide new therapeutic methods for the prevention and treatment of IRI of DCD renal allograft.

心脏死亡捐献（DCD）已成为我国肾移植供体的重要来源，但在等待捐献、获取、保存和移植等过程中发生的缺血再灌注损伤（IRI），却是影响移植肾功能恢复的重要因素。前期研究发现，脂肪基质血管成分（ad-SVF）富含MSC、EPC等干细胞，可有效参与肾脏IRI的修复，具可快速获得，无需体外培养等优点。且肾周ad-SVF还可特异性分泌促红细胞生成素（EPO）等，但供体肾周ad-SVF对心脏死亡来源移植肾IRI的影响目前并不清楚。我们推测：供体肾周ad-SVF可对心脏死亡来源移植肾IRI产生重要的修复作用。本项目拟建立大鼠心脏死亡来源肾移植模型，采用流式细胞分选、microPET-CT示踪及转染等技术，探讨供体肾周ad-SVF对心脏死亡来源移植肾IRI小管上皮及管周毛细血管内皮微环境的影响，阐明MSC、EPC及EPO在供体肾周ad-SVF修复移植肾中的作用机制，可为DCD移植肾IRI的防治提供新途径。

心脏死亡器官捐献（DCD）已成为我国器官来源的重要途径。但DCD患者在等待器官捐献以及供肾获取、保存、移植等过程中发生的移植缺血再灌注损伤（IRI）可导致移植肾功能延迟恢复（DGF）、原发性肾脏无功能等。脂肪基质血管成分（ad-SVF）是机体脂肪组织中的一组不成熟细胞群的总称，表现出比单一的MSC或EPC更强的促进组织器官再生修复的能力，在器官损伤修复与组织再生中显示出了很好的应用潜力。本研究旨在观察ad-SVF对心脏死亡来源的移植肾的保护和修复作用。我们发现ad-SVF可显著减轻肾小管损伤，改善肾功能；同时我们通过流式分选技术，发现CD73+/CD90+/CD45-MSC可通过直接分化促进肾小管上皮细胞修复，CD34+/CD45-EPC可通过直接分化修复移植肾PTC损伤微环境；通过细胞示踪标记，证实供体源性ad-SVF可通过肾动脉注射至肾脏组织，发挥保护和修复作用。免疫组化、western blot、RT-PCR等证明这一保护作用可能与ad-SVF通过自分泌/旁分泌SDF-1、VEGF、EPO等有关。此外，我们通过基因转染等技术，进一步发现SDF-1、VEGF、EPO促进肾小管上皮细胞及内皮细胞的增殖，抑制细胞凋亡，发挥对移植肾的保护和修复作用。结果表明，ad-SVF可促进肾小管上皮和PTC的修复，同时分泌多种活性因子，最终减轻移植肾IRI，改善移植肾肾功能。